Two component signal transduction systems are widely used by pathogenic bacteria to regulate the expression of their virulence factors in response to changing environmental cues during the infectious process. Streptococcus mutans is exposed to the transient environments of the human oral cavity where it is the primary etiologic agent of Dental caries. Specifically, changes in pH, oxygen content, and nutrient availability are likely to necessitate a rapid bacterial response to promote S. mutans-induced cariogenesis. We identified a gene pair in S. mutans that is homologous to the covR/S two-component signal transduction system in the group A streptococci (GAS). CovR/S is known to regulate the expression of multiple genes associated with GAS virulence, including basal which is necessary for capsule production and ska, which encodes a plasminogen activator. Work conducted in5laboratory implicates the S. mutans covR/S homologs in the regulation of fructosyltransferases (ftf) that mediate the sucrose-dependent production of fructans necessary for bacterial adherence to the tooth pellicle. Since S. mutans produces a multitude of factors that promote its survival and persistence in the human host, we posit that CovR/Smay function as a global regulator of S. mutans genes whose products promote disease in the oral cavity. The major goal of this research application is to define a putative role for CovR/S in S. mutans virulence control. The Specific Aims include: 1. Characterization of a S. mutans covR- mutant recently constructed in5laboratory, and analysis of its cariogenic potential in germfree rats; 2. Identification of the S. mutans gene(s) that are subject to CovR/S control in environments that approximate the oral cavity using differential display polymerase chain reaction (ddPCR) and 2D proteomic approaches; 3. construction of S. mutans knockout mutants and reporter gene fusions to functionally characterize CovR/S-regulated genes and analyze their expression. Taken collectively, these studies will elucidate S. mutans mechanism(s) of virulence gene control, and so enable prevention and/or intervention in the pathogenic process that leads to the development of Dental caries.